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Novel Mutations in the katG Gene in Isoniazid Resistant Mycobacterium tuberculosis Isolate Purkan Purkan; Ihsanawati Ihsanawati; Debbie Soefie Soefie; Yana Maolana Syah; Achmad Saifuddin Noer; Dessy Natalia
Jurnal Matematika & Sains Vol 18, No 1 (2013)
Publisher : Institut Teknologi Bandung

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Isoniazid (INH) resistance in Mycobacterium tuberculosis is commonly associated with mutations in katG encoding catalase-peroxidase. Â A clinical isolate of M. tuberculosis L21 exhibits isoniazid resistant phenotype. Â A 2.2 kb DNA fragment of katG gene from the L21 isolate had been cloned and characterized. Â The L21 katG gene has four point mutations, namely T898C; G1388T; T1997C; C2035T; and one G2203 deletion. Â The in silico translation analysis of the katG mutant suggests that the mutation results in a truncated protein lacking of 62 C-terminal amino acids with three amino acid substitutions, Trp300Arg, Arg463Leu, and Ile666Thr. The structural model superposition of the mutant protein with the full length protein suggests that the mutant losses an ability to form a dimer structure, and also posseses a distortion of intermolecular interactions in the substrate binding channel. Keywords: katG gene, KatG, MDR M. tuberculosis, Isoniazid resistance, Structure model. Â Beberapa Mutasi Baru pada Gen katG pada Isolat Mycobacterium tuberculosis Resisten Isoniazid Abstrak Resistensi Mycobacterium tuberculosis terhadap isoniazid umumnya berkaitan dengan mutasi yang terjadi pada gen katG yang pengode katalase-peroksidase.Â Isolat klinis M. tuberculosis L21 fenotip resisten terhadap isoniazid.Â Fragmen DNA berukuran 2,2 kb dari gen katG dari isolate L21 telah diisolasi dan dikarakterisasi. Gen katG dari isolat L21 memiliki empat mutasi, yaitu T898C; G1388T; T1997C; C2035T; dan delesi G2203.Â Analisis in silico terhadap gen katG mutan menunjukkan bahwa mutasi yang terjadi menhasilkan protein mutan yang kehilangan 62 asam amino pada ujung-C yang juga mengandung perubahan asam amino Trp300Arg, Arg463Leu, dan Ile666Thr. Superposisi model struktur protein antara protein mutan dengan protein utuh menyarankan bahwa protein mutan diduga tidak dapat membentuk struktur dimer dan juga mengandung interaksi molekular pada poket pengikatan substrat yang berbeda. Kata kunci: Gen katG, M. tuberculosis, Resisten isoniazid, Model struktur.

ISOLASI DAN KARAKTERISASI BAKTERI PENGHASIL ENZIM LIPASE EKSTRASELULERDARI LUMPUR AKTIF INSTALASI PENGOLAHAN AIR LIMBAH INDUSTRI TEKSTIL Kasipah, Cica; Rismayani, Sinta; Ihsanawati, Ihsanawati; Nurachman, Zeily
Arena Tekstil Vol 28, No 1 (2013)
Publisher : Arena Tekstil

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Lumpur aktif dari instalasi pengolahan air limbah industri tekstil mengandung berbagai jenis mikroorganisme antara lain mikroorganisme yang memiliki aktivitas lipase yang tinggi. Tujuan penelitian ini adalah mengisolasi dan mengkarakterisasi bakteri penghasil enzim lipase ekstraseluler dari lumpur aktif instalasi pengolahan air limbah industri tekstil. Tahapan penelitian yang dilakukan meliputi isolasi bakteri penghasil enzim lipase ekstraseluler dari lumpur aktif melalui skrining dengan media yang mengandung rodamin dan minyak zaitun, penentuan aktivitas enzim lipase dan karakterisasi lipase yang dihasilkan terhadap variasi temperatur, pH, dan pengaruh ion Ca2+. Hasil identifikasi secara mikrobiologi menunjukkan bahwa bakteri penghasil enzim lipase ekstraseluler dari lumpur aktif adalah spesies Erwiniachrysantemi. Enzim lipase ekstraseluler tersebut memiliki aktivitas 4,75 U/mL dengan temperatur optimum 40oC dan pH optimum 9. Penambahan ion Ca2+ tidak memberikan pengaruh berarti terhadap aktivitas enzim lipase.

ISOLASI DAN KARAKTERISASI BAKTERI PENGHASIL ENZIM LIPASE EKSTRASELULERDARI LUMPUR AKTIF INSTALASI PENGOLAHAN AIR LIMBAH INDUSTRI TEKSTIL Cica Kasipah; Sinta Rismayani; Ihsanawati Ihsanawati; Zeily Nurachman
Arena Tekstil Vol 28, No 1 (2013)
Publisher : Balai Besar Tekstil

Lumpur aktif dari instalasi pengolahan air limbah industri tekstil mengandung berbagai jenis mikroorganisme antara lain mikroorganisme yang memiliki aktivitas lipase yang tinggi. Tujuan penelitian ini adalah mengisolasi dan mengkarakterisasi bakteri penghasil enzim lipase ekstraseluler dari lumpur aktif instalasi pengolahan air limbah industri tekstil. Tahapan penelitian yang dilakukan meliputi isolasi bakteri penghasil enzim lipase ekstraseluler dari lumpur aktif melalui skrining dengan media yang mengandung rodamin dan minyak zaitun, penentuan aktivitas enzim lipase dan karakterisasi lipase yang dihasilkan terhadap variasi temperatur, pH, dan pengaruh ion Ca2+. Hasil identifikasi secara mikrobiologi menunjukkan bahwa bakteri penghasil enzim lipase ekstraseluler dari lumpur aktif adalah spesies Erwiniachrysantemi. Enzim lipase ekstraseluler tersebut memiliki aktivitas 4,75 U/mL dengan temperatur optimum 40oC dan pH optimum 9. Penambahan ion Ca2+ tidak memberikan pengaruh berarti terhadap aktivitas enzim lipase.

SKRINING BAKTERI PENDEGRADASI INULIN DARI RIZOSFER UMBI DAHLIA MENGGUNAKAN INULIN UMBI DAHLIA Minda Azhar; Yuni Ahda; Ihsanawati Ihsanawati; Fernita Puspasari; Suci Mawarni; Boni Risa; Dessy Natalia
EKSAKTA: Berkala Ilmiah Bidang MIPA Vol. 18 No. 02 (2017): Eksakta : Berkala Ilmiah Bidang MIPA (E-ISSN : 2549-7464)
Publisher : Faculty of Mathematics and Natural Sciences (FMIPA), Universitas Negeri Padang, Indonesia

Cloning, Overexpression, and Purification of PhoR CytoplasmicDomain Protein from Mycobacterium tuberculosis strain H37Rv OKTIRA ROKA AJI; DYSHELLY NURKARTIKA PASCAPURNAMA; FENRYCO PRATAMA; IHSANAWATI IHSANAWATI; MAELITA RAMDHANI MOEIS; ERNAWATI ARIFIN GIRI-RACHMAN
Microbiology Indonesia Vol. 8 No. 4 (2014): December 2014
Publisher : Indonesian Society for microbiology

Tuberculosis still becomesa major health problem in the world. This infectious disease is caused by Mycobacterium tuberculosis (Mtb). Novel anti-tubercular drug is urgently needed to counter multidrug resistant cases and Mtb's spread. The cytoplasmic domain of PhoR histidine kinase, a part of the two-component system PhoR-PhoP in Mtb, is one of the potential candidates for anti-tubercular drug target.Three dimensional protein of drug target is needed to screen potential drug candidate using rational drug design approaches. Previous studies have successfully characterized and isolated putative cytoplasmic domain of PhoR (CytoPhoR) from Mtb strain H37Rv. This study aimed to clone, overexpress and purify of CytoPhoR protein. CytoPhoR was fused with thioredoxin protein in expression vector pET32b and overexpressed in Escherichia coli (E.coli)BL21 (DE3) as soluble fraction by inductionÂ 1 mM IPTG. Purification of his-tagged CytoPhoR was carried out using IMAC Ni-NTA Agarose his-tag affinity column. SDS-PAGE analysis showed that another protein was co-purified (~35 kDa) along with the CytoPhoR protein. Subsequent protein purification using DEAE-ion exchange column generate a strong single band of 37 kDa on SDSâ€“PAGE which is indicated as CytoPhoR protein. The purified CytoPhoR protein was successfully obtained and can be used for further analysis on determining three dimensional structure of CytoPhoR protein.

Expression and Purification of PhoR Sensor-Domain Histidine Kinase of Mycobacterium tuberculosis in Escherichia coli ERNAWATI ARIFIN GIRI-RACHMAN; FENRYCO PRATAMA; OKTIRA ROKA AJI; ARUM PATRIATI; IHSANAWATI IHSANAWATI; MAELITA RAMDANI MOEIS; EDY GIRI-RACHMAN PUTRA
Microbiology Indonesia Vol. 9 No. 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

Globally, tuberculosis (TB) remains a leading cause of death. The emergence of multidrug-resistant strains (MDR-TB) and extensively drug-resistant strains (XDR-TB) has fuelled the discovery for novel drugs and drug targets for its successful and better treatment. One of the potential candidates for drug target is PhoR sensory protein histidine kinase, a part of the Two Component System (TCS) PhoP/PhoR in Mycobacterium tuberculosis (Mtb). This protein system was known for its role on regulating hundred of Mtb virulence factors, from genes for cell wall and lypid synthesis to genes for adaptation in human leukocyte and hypoxia response. Previous studies have successfully characterized, isolated, and cloned the putative sensory domain of PhoR protein gene into pRSET vector expression system. In this study, Escherichia coli was transformed with pRSET-SensPhoR and cultivated at 37oC under IPTG induction to express PhoR sensor-domain protein. Most of the proteins were overexpressed in the form of inclusion bodies. Â Subsequent protein purification in Ni-NTA system under refolding condition on urea gradient was performed to isolate PhoR sensor-domain protein in soluble form. Arginine was supplemented in purified protein solution to prevent aggregation during long term storage. Â While highly purified protein was acquired, small angle X-ray scattering (SAXS) analysis was conducted to obtain 3-dimensional (3D) protein structures in solution. Â Â Â doi:10.5454/mi.9.2.1Â

Antibacterial Activity of Endophytic Fungus Isolates of Mangrove Fruit (Sonneratia alba) Against Staphylococcus aureus and Esherichia coli Sesilia Rante Pakadang; Ismat Marsus; Ihsanawati Ihsanawati
JURNAL INFO KESEHATAN Vol 19 No 1 (2021): JURNAL INFO KESEHATAN
Publisher : Research and Community Service Unit, Politeknik Kesehatan Kemenkes Kupang

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.31965/infokes.Vol19.Iss1.416

Endophytic fungi live and associate in plant tissues and have a mutualistic relationship. Endophytic fungi produce various compounds such as steroids, terpenoids, phenolics, alkaloids which are the same as secondary metabolites from their host plants. The objective of this study is to isolate and identify endophytic fungi from mangrove fruit (Sonneratia alba) and to determine the antibacterial activity of endophytic fungi isolates against the growth of Staphylococcus aureus and Escherichia coli. The type of research is pre-experimental design, one shoot case study. The methods used are isolation, identification and agar diffusion. The pieces of mangrove fruit were disinfected and then cultured on SDA media to grow endophytic fungi isolates. The isolates were cultured repeatedly until pure isolates were obtained. The test of isolate activity against antibacterial was determined by the agar diffusion method with the test material of 2 isolates of endophytic fungi on Nutrient Agar (NA) media. The results showed that the mangrove fruit (Sonneratia alba) produced two isolates of endophytic fungi that could inhibit the growth of Staphylococcus aureus and Escherichia coli. It was concluded that the mangrove fruit culture produced 2 isolates, which are isolate 1 Aspergillus niger and isolate 2 Aspergillus flavus. Isolate 1 and isolate 2 had the potential as antibacterial against the growth of Staphylococcus aureus and Escherichia coli. Isolate 2 was more effective than isolate 1 in inhibiting the growth of Staphylococcus aureus. It is recommended to test the pharmacological and microbiological activity of the findings of isolates 1 and 2 in vivo.

New Pyrrole-Chalcone Hybrids Against Acetylcholinesterase: Synthesis, In Vitro, and Computational Studies Ridwanuloh, Dadan; Danova, Ade; Hermawati, Elvira; Chavasiri, Warinthorn; Ihsanawati, Ihsanawati; Alni, Anita
Journal of Multidisciplinary Applied Natural Science Articles in Press
Publisher : Pandawa Institute

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.47352/jmans.2774-3047.375

Acetylcholinesterase (AChE) inhibition remains a central therapeutic approach for Alzheimer’s disease (AD), as it helps preserve synaptic acetylcholine levels, enhances cholinergic neurotransmission, and mitigates early cognitive decline. In this study, eight novel pyrrole–chalcone hybrids (3–10), consisting of four pyrrole–chalcones (3–6) and four pyrrole–chalcone amides (7–10), were designed, synthesized, and biologically evaluated for AChE inhibition. Among them, compound 8 (N-(4-methoxybenzyl)-pyrrole–chalcone amide) and compound 10 (N-(3,4-dimethoxybenzyl)-pyrrole–chalcone amide) demonstrated the strongest inhibitory activity, with IC₅₀ values of 3.1 and 2.8 µM, respectively, comparable to galantamine. Kinetic assays confirmed that both compounds act as noncompetitive inhibitors, as indicated by reduced Vmax without significant alteration in Km, while compound 10 exhibited Ki of 0.8 µM, reflecting high enzyme affinity. Molecular docking revealed strong binding interactions of compounds 8 and 10 with key AChE residues (Trp84, Phe330, Tyr334), supported by π–π stacking, π–alkyl interactions, and hydrogen bonding, with binding energies of –9.2 (compound 8) and –8.9 kcal/mol (compound 10). Molecular dynamics simulations further demonstrated that compound 10 forms a more stable and compact complex with AChE, as indicated by consistent RMSD values and a stable radius of gyration. SwissADME analysis confirmed favorable pharmacokinetic profiles for both ligands, including Lipinski compliance, high GI absorption, and absence of PAINS alerts, despite the lack of predicted BBB permeability. Overall, compound 10 emerges as the most promising noncompetitive AChE inhibitor in this series, exhibiting strong binding affinity, structural stability, and drug-likeness, thus warranting further optimization and in vivo evaluation.

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